Perhaps one of the most influential concepts in protein mass spectrometry has been the notion of enzymatic protein digestion to render a collection of peptides of suitable size for conventional tandem mass spectrometry (collisional-activation, CAD).1 Doubtless this methodology has enabled significant progress for global protein identification;however, many investigators now realize this approach has significant limitations.2 This conclusion is based upon the following observations: First, protein posttranslational modifications (PTMs) on multi-domain proteins, and among components of protein-protein machines, work in concert;to determine their biological relevance, these patterns must be detected within the context of one another (across the whole protein).3 Second, transcriptional editing processes are pervasive in higher eukaryotes and difficult to predict, even with a completely sequenced genome. For example, 3/4 of all human proteins are expected to have at least 1 splice variant4-6 - variants that could contain intronic sequences. Skipped codons, frameshifting, gene fusion, and single nucleotide polymorphisms (SNPs) also occur. Thus, the use of short peptides as proxy markers for genes is inadequate and often misleading. Unlike CAD, electron transfer dissociation (ETD), a new fragmentation technique co-invented by the PI, does not require short peptides for successful sequence analysis (i.e., trypsin digestion). ETD is indifferent to peptide length or the presence of PTMs, is performed on a time-scale that permits coupling with chromatography, and can be coupled to other ion/ion reactions. This proposal aims to develop a suite of core ion/ion reaction tools, and automate their use in a hybrid-